The Tension-Activated Channels In The Cytoplasmic Membrane Of Vibrio Cholerae

Pathogenic Vibrio cholerae easily transitions between fresh water reservoirs and host intestines, having exceptional adaptation to abrupt osmotic changes. Mechanosensitive (MS) channels in the inner membrane act as tension-sensitive release valves, ejecting osmolytes with water during osmotic downshock, allowing the bacteria to maintain an optimal turgor within the cell and avoid lysis. We report the absolute tensions associated with the opening of MscS- and MscL-like channels (7.3 and ∼11 mN/m, respectively), determined by imaging and simultaneous patch-clamping V. cholerae spheroplasts in whole-cell mode. By cloning the MS channel homologs from V. cholerae and studying them in E. coli spheroplasts we determined that the major contributors to these mechanoelectrical responses are VcMscS, a 287 aa protein 52% identical to EcMscS, a ‘long’ 569 aa homolog VcMscS-L (36% identical to EcMscS), and a single VcMscL (136 aa, 65% identity to EcMscL). We found that the lateral tension in the membrane, which drives gating of these channels, can be modulated by asymmetric stress induced by intercalation of exogenous amphipathic substances. CAI-1, a quorum sensing molecule involved in signaling biofilm formation in stressed Vibrio populations has strong amphipathicity. Whether CAI-1 permeates through the membrane directly or requires specific transporters remains unknown. In response to asymmetric addition of CAI-1 we observed shifts of activation curves of V. cholerae MS channels toward higher pressure. Measurements of lateral pressures created by different concentrations of CAI-1 in native membranes and in lipid monolayers gave estimations of the membrane association constant as ∼106 M-1. This suggests that CAI-1 does not require transporters and permeates through the membrane directly. Thus, tension-activated channels which act as osmolyte release valves in the cytoplasmic membrane of Vibrio cholerae can be used as lateral pressure gauges to detect partitioning of lipophilic signaling molecules.